摘要
为了创制大白菜—结球甘蓝异附加系,以大白菜—结球甘蓝异源三倍体杂交种(AAC,2n=3x=29)与二倍体大白菜(AA,2n=2x=20)回交产生的BC_1代及其自交后代BC_1F_2为试验材料,采用常规压片、核型分析,并结合SSR分子标记方法进行了鉴定,获得了附加甘蓝染色体的2个单体异附加系、1个易位系、1个双体异附加系,以及2个双单体异附加系,并对鉴定出的异附加系进行了形态学、细胞学和生殖特性方面的研究。本研究为异代换系和易位系的构建提供了基础材料,也为研究A、C基因组的亲缘关系、基因定位和表达以及加速大白菜品种遗传改良奠定了基础。研究结果如下:
1.通过对BC_1代植株进行根尖有丝分裂观察,BC_1F_2代植株花粉母细胞减数分裂后期Ⅰ观察,对BC_1和BC_1F_2各个单株染色体数目进行了鉴定。98个BC_1代植株中染色体数为21和22条的各占4.08%、5.10%,16个BC_1F_2代植株中染色体数为21和22条的均占12.5%。两个世代均以染色体数为23～24植株占多数,分别占55.1%、31.25%。
2.有相同染色体数目的BC_1F_2植株田间性状表现出多样性。附加外源染色体的植株在某些方面有相似于结球甘蓝的形态特征,这有助于根据植株田间性状进行附加染色体类别的鉴定。
3.采用常规压片技术对BC_1和BC_1F_2的染色体数进行了统计,通过核型分析初步鉴定出附加结球甘蓝2号和4号染色体的3个单体异附加系,附加2号染色体的1个双体异附加系,以及两个分别附加2号、4号和4号、9号的双单体异附加系。对鉴定出的异附加系植株进行了田间性状观察,发现与亲本大白菜相比均出现类似结球甘蓝的不同性状,且附加不同甘蓝染色体,其表现具有明显差异。附加结球甘蓝4号染色体的两个株系植株性状明显不同,因此对其进行SSR分子标记鉴定发现,其中一个株系染色体可能发生易位。
4.通过对鉴定出的植株进行花粉母细胞减数分裂行为观察发现:(1)染色体数为21的3个株系AA+C_(2Ⅱ),AA+C_(4Ⅰ),AA+C_(4T)减数分裂终变期大多以10Ⅱ+Ⅰ方式联会,极少数以9Ⅱ+Ⅲ方式联会,后期Ⅰ植株染色体基本以11-10的方式分离,后期Ⅱ时,多以11-10-11-10的方式分离,少数细胞有一个或两个落后染色体出现。(2)染色体数为22条的双体异附加系AA+C_(2D)终变期以11Ⅱ方式联会,中期Ⅰ染色体均匀排列于赤道板,后期Ⅰ由于落后染色体导致多数细胞以10-2-10的方式分离,少数细胞以11-11方式分离。(3)染色体数为22条的两个双单体异附加系株系w42、e45终变期均以10Ⅱ+Ⅰ+Ⅰ的方式联会,后期Ⅰ以12-10、11-11、10-2-10多种方式分离。(4)染色体数大于22条的植株终变期染色体出现多种联会的形式,出现多价体的频率大幅增加,导致中后期有较多的染色体落后或丢失,分离形式表现出多样性。
In order to create the alien addition lines, the BC_1 plants devived from the backcrosse between the Chinese cabbage-cabbage allotriploid (ACC,2n=3x=29) and diploid Chinese cabbage (AA, 2n=2x=20) and their selfing progenies BC_1F_2 plants are used as studying materials. Two monosomic addition lines, one translocation line, one disomic addition line and two double alien monosomic addition lines are obtained by routine planishing technique , karyotype analysis and SSR molecular marks. And their morphologic, cytologic and reproductive characters are studied. This study provids basic material for the alien substitution lines and the translocation lines and also help to research the relative of A and C genomes, gene mapping and expression and the genetic improvement of varietices. The main results are showed as follows:
1. The chromosome number of BC_1 was identified by counting chromosomes of tipced at mitosis, and the chromosome number of BC_1F_2 was identified by counting chromosomes of PMC at meiosis anaphase I. The rate of plants with 21 chromosomes and 22 chromosomes identified from 98 plants of BC_1 and 16 plants of BC_1F_2 was 4.08% and 5.10%, 12.5% and 12.5% respectively. The chromosome number of most plants was 23 to 24,with rate 55.10% in BC_1 and 31.25% in BC_1F_2.
2. The field traits of BC1F2 plants with the same chromosome number showed multiplicity. The plants with extra chromosomes seemed like the parent cabbage in certain aspects, which would help to identify the type of addition chromosome according to field characters.
3. Three monosomic addition lines with cabbage chromosome 2 and 4 , one disomic alien addition line with cabbage chromosome 2 and two double alien monosomic addition lines with cabbage chromosome2,4 and cabbage chromosome 4,9 were obtained by karyotype analysis. The field characters of the plants obtained were observed. The results showed that the alien addition lines seemed like the parent cabbage in certain aspects comparing with the diploid parent Chinese cabbage. And the characters of the plants with different cabbage chromosomes were very different. The field characters of two plants both with cabbage chromosome 4 were very different, and the results from SSR molecular marks of them showed that one of them may be a translocation line.
4.The PMC meiosis behavior of the plants obtained showed that: 1)To the plants with 21 chromosomes AA+C_(2II),AA+C_(4I),AA+C_(4T), the chromosome configuration at diakinesis was mainly 10II+I, few of them were 9II+III. The form_I of chromosome separation at anaphase I was mainly 11-10, and the form of chromosome separation at anaphase II was mainly 11-10-11-10,one or two lagging chromosomes appeared in few cells. 2) To the disomic alien addition lines with 22 chromosomes AA+C_(2D), the chromosome configuration at diakinesis was 11II; chromosomes moved to the equator plank at metaphase I ; the form of chromosome separation at anaphase I was mainly 10-2-10,and the form 11-11 occurred in few cells because of the lagging chromosomes. 3) To the double alien monosomic addition lines with 22 chromosomes w42 and e45, the chromosome configuration at diakinesis was mainly 10II+I+I, and multiple forms of chromosome separation at anaphase I, including 11-11,12-10 and 10-2-10 were deserved. 4) To the plants with more than 22 chromosomes, there were many formations of chromosome synapsis at diakinesis. The rate of multivalents was increased distinctly, which led to more lagging or losing chromosomes. The above separation formation indicated the multiformity.
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